Polymer hydrogels have structural similarities to numerous macromolecular components in the human body, are generally considered biocompatible, and have been investigated extensively as materials useful for drug delivery, tissue repair and tissue engineering, as well as for use as surgical sealants and adhesives. With increasing frequency, polymer hydrogels are designed for in situ gelation from a liquid precursor, thereby allowing minimally invasive administration via syringe or needle.
Existing hydrogel systems, formed chemically or physically, are subject to several limitations. Chemically cross-linked hydrogels often employ toxic cross-linking agents and/or radicals, and the resulting hydrogels are often non-biodegradable. On the other hand, physical hydrogels—formed through ionic interactions, hydrophobic interactions, hydrogen bonding, or phase transition—are relatively weak and can be prone to unwanted or uncontrollable degradation through ion exchange, ion diffusion, or monomer dissolution. An alternate approach—solidification by enzymatic cross-linking—has two principal advantages, compared to other hydrogel systems. First, an enzyme has substrate specificity to allow controllable gel formation. Second, an enzymatic method can be applied to the in vivo utilization of cross-linked hydrogels, under appropriate physiological conditions.
Transglutaminases (e.g. protein-glutamine:amine γ-glutamyltransferase, EC 2.3.2.13) catalyze a post-translational acyl-transfer reaction between the γ-carboxamide groups of peptide-bound glutamine residues and the ε-amino groups of lysine residues in proteins, or certain primary amino groups, resulting in the cross-linking of proteins through the formation of ε-(γ-glutamyl)lysine isopeptide side-chain bridges. Although several biological fluids are known to undergo rapid transglutaminase (TGase) catalyzed hydrogel formation, previous attempts to use TGase with peptide modified synthetic polymers have resulted in slowly gelling systems. Although proteins are typical TGase substrates, the prior art has demonstrated that synthetic poly(amino acids) and peptide-modified poly(ethylene glycol) can be cross-linked with hydrogel formation. Recently, factor XIII (plasma transglutaminase) was reported to catalyze a hydrogel formation. However, due to the stringent substrate specificity of factor XIII, a 20 amino acid-long peptide from the cross-linking γ-chain of fibrinogen was first, necessarily, synthesized and conjugated to a branched polyethylene glycol (PEG) polymer. The complex preparation of the initial conjugate made a large-scale hydrogel preparation difficult. In both this instance and using synthetic peptides, the time required for gelation was considerably longer than useful for many clinical applications.